Analysis of oncogene and microRNA involvement in neuroblastoma chemosensitivity modulation

Abstract

ABSTRACT:Neuroblastoma (NB) is a paediatric tumour arising from the neural crest cells, whichtypically differentiate into neural cells of the sympathetic nervous system. Prognosis ofNB is dire, with less than 40 % of all paediatric patients surviving, following diagnosiswith aggressive NB tumours. First line treatment for children afflicted with advancedneuroblastoma activity usually involves chemotherapy. However, the main problemswith the use of chemotherapeutic agents are the dose-dependent adverse effects andthe risk of development of cytotoxic drug resistant tumours. The purpose of this studywas to identify dysregulations in expression of individual and / or networks of microRNAs (miRNAs) that may have direct effect on neuroblastoma (NB) drug resistance.Furthermore, analysis of the downstream effect of recognised NB oncogene expressionon genes directly linked to multi-drug resistance (MDR) was evaluated accordingly.METHODS: Validated NB chemoresistance models containing the chemosentitiveparental and subcultured chemoresistant cell lines were analysed for miRNAexpression profiling, using a high-throughput quantitative polymerase chain reaction(RT-qPCR) miRNA profiling platform for a total of 668 miRNAs. Validation of anyidentified putative chemoresistance miRNAs by RT-qPCR, and chemoresistancefunctional analyses through transient and stable transfection of miRNA antagonist /mimic molecules were also performed. RT-qPCR was also performed on tworecognised oncogenes (MYCN, ALK) and six individual MDR genes within two NBcell lines, namely SH-EP1 and LA1N55n.RESULTS:A total of two putative chemoresistance miRNAs, hsa-miR-125b-1# andhsa-miR-188-5p were validated up to the level of stable transfection with miRNAantagonists. However, the influence of individual miRNA modulation on NB cell linechemoresistance phenotype was not elucidated. No statistically significantdysregulated expression of the studied MDR genes was identified, within MYCNamplified NB cell lines.CONCLUSIONS: Based on the initial miRNA findings, this study elucidates theimportance of multi-factorial elements which contribute dynamically and inorchestration for the attainment and shifting of complex phenotype characteristics suchas the capacity of the individualcell to withstand exposure to cytotoxicchemotherapeutic agents commonly utilised in the clinic for neuroblastoma patients.

Date of Award	1 Jan 1824
Original language	English
Awarding Institution	The University of Manchester
Supervisor	William Ollier (Supervisor) & Philip Day (Supervisor)